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4, 150?mM NaCl, 1?mM EDTA, 1% Triton X-100, 1% Sodium deoxycholate, 0.1% SDS and proteinase inhibitor). The protein was removed from the beads by boiling for 5?min in 1X LSB (2% SDS, 10% glycerol, 6.25?mM Tris pH 6.8). Proteins were diglyceride separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes. Membranes were blocked in 5% nonfat dry milk and then probed with 0.4?��g/ml goat polyclonal antibody against mouse BSG (R&D) overnight at 4?��C. The membranes were then washed and incubated with horseradish peroxidase-conjugated donkey anti-goat IgG antibody (abcam, San Francisco, CA) at 1:5000 dilution for 60?min at room temperature.
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